Trypanosoma cruzi is an important human pathogen responsible for 10-50,000 deaths/year infecting 12-20 million people. Vector control measures have met with some success but available drug therapies have low efficacy and high toxicity, and no specific vaccines have entered human trials. There is a desperate need for a safe and effective vaccine to protect the 40-100 million individuals at risk of acquiring this deadly diseas. Robust CD8+ T cell responses are detected against parasite lysate and a variety of T. cruzi proteins during chronic infection. In fact, increased frequencies of T. cruzi-specific IFN-? producing CD8+ T cells correlate with decreased disease progression. We have developed animal models to carefully analyze vaccine-induced immune responses and cardiac pathology associated with T. cruzi infection. Several studies have shown that CD4+ T cells are necessary during priming functions of protective immunity, and CD8+ T cells are essential for effector function and parasite clearance. T. cruzi trans-sialidase (TS) antigens can induce potent murine protection against normally lethal systemic and sub-lethal mucosal T. cruzi challenges; however, TS vaccines have not been optimized for human use. In addition, the large TS gene family represents >5% of all T. cruzi genes, and expression of some TS antigens may be involved in parasite immunoevasion. We will address this latter possibility by comparing vaccines encoding epitopes from functional TS genes (encoding the 12-15 enzymatically active proteins), non-functional TS genes (>700 genes), and mammalian stage non-TS genes. We propose to identify key parasite epitopes from these important T. cruzi protein sets predicted to bind common HLA alleles (providing selection coverage relevant for >95% of all human populations) utilizing state- of-the-art immunoinformatics. Predicted epitope sequences will be synthesized and HLA binding assays will be employed to validate MHC binding. In collaboration with Dr. Meymandi at UCLA, we will obtain blood samples from individuals infected with T. cruzi (and uninfected controls) and stimulate purified PBMC with peptides described above in IFN-?? ELISPOT and flow cytometric assays. Based on results from these screening assays, we will design novel vaccines encoding these T cell epitopes, and test their prophylactic and therapeutic efficacies in 'humanized' HLA A2/DR1 dual transgenic mice. The outcome of our studies will provide key data required for proceeding with phase I Chagas vaccine trials. PUBLIC HEALTH RELEVANCE: Approximately 12-20 million people are infected with T. cruzi, 10-50,000 people die each year from Chagas disease, people in the US are at risk of T. cruzi infection through blood transfusions and organ donations originating from infected persons, and there are no highly effective vaccines or chemotherapies for the prevention or treatment of T. cruzi infection and disease. The work proposed in this application focuses on the identification of T. cruzi T cell epitopes and will generate and test novel vaccines in 'humanized' HLA transgenic mice. Results of our studies will provide important new information relevant for the development of more effective T. cruzi vaccines, as well as the development of vaccines for other major human diseases (e.g.-AIDS, TB) caused by mucosally invasive, intracellular infections.